Haddad I Y, Pataki G, Hu P, Galliani C, Beckman J S, Matalon S
Department of Pediatrics, University of Alabama at Birmingham 35233-6810.
J Clin Invest. 1994 Dec;94(6):2407-13. doi: 10.1172/JCI117607.
Activated alveolar macrophages and epithelial type II cells release both nitric oxide and superoxide which react at near diffusion-limited rate (6.7 x 10(9) M-1s-1) to form peroxynitrite, a potent oxidant capable of damaging the alveolar epithelium and pulmonary surfactant. Peroxynitrite, but not nitric oxide or superoxide, readily nitrates phenolic rings including tyrosine. We quantified the presence of nitrotyrosine in the lungs of patients with the adult respiratory distress syndrome (ARDS) and in the lungs of rats exposed to hyperoxia (100% O2 for 60 h) using quantitative immunofluorescence. Fresh frozen or paraffin-embedded lung sections were incubated with a polyclonal antibody to nitrotyrosine, followed by goat anti-rabbit IgG coupled to rhodamine. Sections from patients with ARDS (n = 5), or from rats exposed to hyperoxia (n = 4), exhibited a twofold increase of specific binding over controls. This binding was blocked by the addition of an excess amount of nitrotyrosine and was absent when the nitrotyrosine antibody was replaced with nonimmune IgG. In additional experiments we demonstrated nitrotyrosine formation in rat lung sections incubated in vitro with peroxynitrite, but not nitric oxide or reactive oxygen species. These data suggest that toxic levels of peroxynitrite may be formed in the lungs of patients with acute lung injury.
活化的肺泡巨噬细胞和II型上皮细胞会释放一氧化氮和超氧化物，它们以接近扩散极限的速率（6.7×10⁹ M⁻¹s⁻¹）发生反应，形成过氧亚硝酸盐，这是一种能够损伤肺泡上皮和肺表面活性物质的强效氧化剂。过氧亚硝酸盐而非一氧化氮或超氧化物，能够轻易地使包括酪氨酸在内的酚环硝化。我们使用定量免疫荧光法，对成人呼吸窘迫综合征（ARDS）患者的肺组织以及暴露于高氧环境（100%氧气，持续60小时）的大鼠肺组织中硝基酪氨酸的存在情况进行了定量分析。将新鲜冷冻或石蜡包埋的肺组织切片与抗硝基酪氨酸的多克隆抗体孵育，随后与偶联有罗丹明的山羊抗兔IgG孵育。ARDS患者（n = 5）或暴露于高氧环境的大鼠（n = 4）的切片显示，特异性结合较对照组增加了两倍。加入过量的硝基酪氨酸可阻断这种结合，当用非免疫IgG替代硝基酪氨酸抗体时则不存在这种结合。在其他实验中，我们证明在体外与过氧亚硝酸盐而非一氧化氮或活性氧一起孵育的大鼠肺组织切片中会形成硝基酪氨酸。这些数据表明，急性肺损伤患者的肺组织中可能会形成毒性水平的过氧亚硝酸盐。
